N2pc reflects two modes for coding the number of visual targets.

Humans share with a variety of animal species the spontaneous ability to detect the numerical correspondence between limited quantities of visual objects and discrete auditory events. Here, we explored how such mental representation is generated in the visual modality by monitoring a parieto-occipital ERP component, N2pc, whose amplitude covaries with the number of visual targets in explicit enumeration. Participants listened to an auditory sequence of one to three tones followed by a visual search display containing one to three targets. In Experiment 1, participants were asked to respond based on the numerical correspondence between tones and visual targets. In Experiment 2, participants were asked to ignore the tones and detect a target presence in the search display. The results of Experiment 1 showed an N2pc amplitude increase determined by the number of visual targets followed by a centroparietal ERP component modulated by the numerical correspondence between tones and visual targets. The results of Experiment 2 did not show an N2pc amplitude increase as a function of the number of visual targets. However, the numerical correspondence between tones and visual targets influenced N2pc amplitude. By comparing a subset of amplitude/latency parameters between Experiment 1 and 2, the present results suggest N2pc reflects two modes for representing the number of visual targets. One mode, susceptible to subjective control, relies on visual target segregation for exact target individuation, whereas a different mode, likely enabling spontaneous cross-modal matching, relies on the extraction of rough information about number of targets from visual input.

Impact of novel histopathological factors on the outcomes of liver surgery for colorectal cancer metastases.

INTRODUCTION: We evaluated the impacts of a series of novel histopathological factors on clinical-surgical outcomes and survival of patients who underwent surgery for colorectal cancer liver metastasis, with and without neoadjuvant chemotherapy. MATERIALS AND METHODS: A prospective database including 150 consecutive patients who underwent 183 hepatic resections for metastatic colorectal cancer was evaluated. Among them, 74 (49.3%) received neoadjuvant chemotherapy before surgery. The histopathological factors studied were: a) microsatellitosis, b) type and pattern of tumour growth, c) nuclear grade and the number of mitoses/mm(2), d) perilesional pseudocapsule, e) intratumoural fibrosis, f) lesion cellularity, g) hypoxic-angiogenic perilesional growth pattern, and h) the tumour normal interface. RESULTS: Three or more metastatic lesions, R1 resection margins, and <50% tumour necrosis were prognostic factors for a worse OS, but only the former was confirmed to be an independent prognostic factor in the multivariate analysis. Furthermore, tumour fibrosis <40% and cellularity >10% were predictive of a worse neoadjuvant therapy response, but these findings were not confirmed in the multivariate analysis. Finally, tumour necrosis <50%, cellularity >10%, and TNI >0.5 mm were prognostic factors for a worse DFS and AS in the univariate but not in the multivariate analysis. CONCLUSIONS: Several factors seem to influence the outcomes of surgery for colorectal cancer liver metastasis, especially the number of the lesions, the margins of resection, the percentage of necrosis and fibrosis, as well as the cellularity and the TNI.

Functional significance of the novel H-RAS gene mutation M72I in a patient with medullary thyroid cancer.

Medullary thyroid cancer (MTC) accounts for around 5-10% of all thyroid cancers. Though usually sporadic, 1 in 4 cases are of genetic origin, with germinal mutations in the RET proto-oncogene in familial forms and somatic mutations both in RET and in the RAS family genes in sporadic ones.This study aimed to characterize a rare H-RAS sequence variant -M72I- in a patient with sporadic MTC, focusing on its functional significance.Mutation analysis was performed for the RET, N-RAS, K-RAS and H-RAS genes by direct sequencing. Western blot analysis was done on 4 thyroid tissues from 1 patient carrying the M72I mutation in H-RAS, 1 with the Q61R mutation in H-RAS, 1 with no RET, H-RAS, K-RAS or N-RAS gene mutations, and 1 normal thyroid, using different antibodies against Erk1/2, phospho-Erk1/2 (Thr202/Tyr204), Akt and phospho-Akt (Ser473). Large-scale molecular dynamics simulations were completed for H-RAS wt and H-RAS M72I.Western blot analysis demonstrated that both MAPK and PI3K/Akt pathways were activated in the MTC patient carrying the M72I variant. In silico results showed conformational changes in H-RAS that could influence its activation by Sos and phosphate binding. Results of molecular dynamics were consistent with Western blot experiments.The M72I mutation may contribute effectively to proliferation and survival signaling throughout the MAPK and PI3K/Akt pathways. This work underscores the importance of studying genetic alterations that may lead to carcinogenesis.

ß2 -Glycoprotein I binds to thrombin and selectively inhibits the enzyme procoagulant functions.

BACKGROUND: This work was aimed at characterizing the interaction of ß(2)-glycoprotein I (ß(2)GPI), an abundant plasma protein of unknown function, with human thrombin, the final effector protease in the coagulation cascade. METHODS: The ß(2)GPI-thrombin interaction was studied by surface plasmon resonance (SPR), fluorescence, and molecular modeling. The effect of ß(2)GPI on the procoagulant (fibrin generation and platelet aggregation) and anticoagulant (protein C activation) functions of thrombin were investigated with turbidimetric, immunocytofluorimetric and enzymatic assays. RESULTS: SPR and fluorescence data indicated that ß(2)GPI tightly bound thrombin (K(d) = 34 nM) by interacting with both protease exosites, while leaving the active site accessible. This picture is fully consistent with the theoretical model of the ß(2)GPI-thrombin complex. In particular, blockage of thrombin exosites with binders specific for exosite-1 (hirugen and HD1 aptamer) or exosite-2 (fibrinogen γ'-peptide and HD22 aptamer) impaired the ß2 GPI-thrombin interaction. Identical results were obtained with thrombin mutants having one of the two exosites selectively compromised by mutation (Arg73Ala and Arg101Ala). Fluorescence measurements indicated that ß(2)GPI did not affect the affinity of the enzyme for active site inhibitors, such as p-aminobenzamidine and the hirudin(1-47) domain, in agreement with the structural model. ß(2)GPI dose-dependently prolonged the thrombin clotting time and ecarin clotting time in ß(2)GPI-deficient plasma. ß(2)GPI inhibited thrombin-induced platelet aggregation (IC50 = 0.36 µM) by impairing thrombin cleavage of protease-activated receptor 1 (PAR1) (IC50 = 0.32 µM), both on gel-filtered platelets and in whole blood. Strikingly, ß(2) GPI did not affect thrombin-mediated generation of the anticoagulant protein C. CONCLUSIONS: ß(2) GPI functions as a physiologic anticoagulant by inhibiting the key procoagulant activities of thrombin without affecting its unique anticoagulant function.

Kinase CK2 inhibition: an update.

Protein kinase CK2 (Casein Kinase 2) is an essential, ubiquitous and highly pleiotropic protein kinase, implicated in several human diseases. In the last decade, several inhibitors of CK2, have been discovered and characterized to be ATP-competitive compounds. However, only one of them, CX-4945, has recently completed Phase I clinical trial as potential anticancer drug. In this review, we report all chemical classes of CK2 inhibitors available in literature, focusing our attention on conventional ATP-competitive and on non ATP-competitive inhibitors, which could represent a new frontier in CK2 inhibition and, consequently, a promising field of study in discovering new drug candidates.

The dark side of protein kinase CK2 inhibition.

Casein kinase 2 (CK2) is a ubiquitous, highly pleiotropic and essential protein kinase whose abnormally high constitutive activity has been implicated in several human diseases. In the last decade, several ATP competitive inhibitors of CK2, characterized by an in vitro activity that ranges from micromolar to nanomolar, have been discovered. However, until now only one drug candidate has been entered in Phase I clinical trial as a potential anticancer drug. Why this constitutively active kinase is so undruggable? Can ATP competitive inhibitors be considered the most promising drug candidates for the near future? In this review, we would like to underline how targeting binding sites outside the conventional ATP-binding could represent a new promising strategy to inhibit CK2 activity and, consequently, bear a great potentiality in discovering new drug candidates.

Bandwidth-efficient phase modulation techniques for stimulated Brillouin scattering suppression in fiber optic parametric amplifiers.

Two novel bandwidth efficient pump-dithering Stimulated Brillouin Scattering (SBS) suppression techniques are introduced. The techniques employ a frequency-hopped chirp and an RF noise source to impart phase modulation on the pumps of a two pump Fiber Optical Parametric Amplifier (FOPA). The effectiveness of the introduced techniques is confirmed by measurements of the SBS threshold increase and the associated improvements relative to the current state of the art. Additionally, the effect on the idler signal integrity is presented as measured following amplification from a two pump FOPA employing both techniques. The measured 0.8 dB penalty with pumps dithered by an RF noise source, after accruing 160 ps/nm of dispersion with 38 dB conversion gain in a two-pump FOPA is the lowest reported to date.

Short wavelength infrared frequency conversion in ultra-compact fiber device.

Linear and nonlinear characteristics of devices using millimeter-scale spools of highly nonlinear fiber are experimentally investigated within 2000-2400nm spectral range. Coils with radius larger than 3.5 mm indicate that macro-bending induced radiation loss is negligible up to 2400nm. Devices with smaller diameter coiling resulted in macro-bending losses that dominate over micro-bending losses beyond 2200nm. A tunable short-wave infrared source was constructed using a coin-sized fiber module to demonstrate an efficient nonlinear conversion from 1.26 to 2.2 microm.

DNA topoisomerase II structures and anthracycline activity: insights into ternary complex formation.

DNA Topoisomerase II (Top2) is an essential nuclear enzyme that regulates the topological state of the DNA, and a target of very effective anticancer drugs including anthracycline antibiotics. Even though several aspects of drug activity against Top2 are understood, the drug receptor site is not yet known. Several Top2 mutants have altered drug sensitivity and have provided information of structural features determining drug action. Here, we have revised the published crystal structures of eukaryotic and prokaryotic Top2s and relevant biochemical investigations of enzyme activity and anthracycline action. In particular, we have considered Top2 mutations conferring resistance to anthracyclines and related agents. Following a previous study (Moro et al, Biochemistry, 2004; 43: 7503-13), we have then re-built a molecular model of the entire enzyme in complex with DNA after the cleavage reaction, and used it to define the receptor site of anthracyclines. The results suggest a model wherein the drug specifically contacts the cleaved DNA as well as amino acid residues of the enzyme CAP-like domain. The findings can explain several established structure-activity relationships of antitumour anthracyclines, and provide a framework for further developments of effective Top2 poison.

Tandem 3D-QSARs approach as a valuable tool to predict binding affinity data: design of new Gly/NMDA receptor antagonists as a key study.

Quantitative structure-activity relationships (QSARs) represent a very well consolidated computational approach to correlate structural or property descriptors of chemical compounds with their chemical or biological activities. We have recently reported that autocorrelation Molecular Electrostatic Potential (autoMEP) vectors in combination to Partial Least-Square (PLS) analysis or to Response Surface Analysis (RSA) can represent an interesting alternative 3D-QSAR strategy. In the present paper, we would like to present how the applicability of in tandem linear and nonlinear 3D-QSAR methods (autoMEP/PLS&RSA) can help to predict binding affinity data of a new set of N-methyl-d-aspartate (Gly/NMDA) receptor antagonists.

Recovery of vision and pupil responses in optic neuritis and multiple sclerosis.

The recovery of visual performance and pupil responses were investigated in patients with demyelinating optic neuritis (ON) and multiple sclerosis (MS). The pupil constriction amplitude and the time delay (latency) of the pupil response were measured in 14 patients with a history of unilateral ON in response to either achromatic (luminance) or chromatic (isoluminant) stimulus modulation. Five of these subjects were diagnosed later with MS. In addition, we measured detection thresholds for achromatic stimuli using standard visual field perimetry and chromatic thresholds using a new colour assessment and diagnosis (CAD) test that isolates the use of colour signals. The results show that, despite significant improvements in visual function following the acute phase (as assessed using visual acuity and fields), significant pupil response deficits remain. The findings also demonstrate that accurate measurements of pupil responses and chromatic thresholds can reveal deficits that remain undetected with more conventional techniques. These preliminary findings suggest that the techniques described here can provide useful information about remitting and relapsing demyelinative phases, often observed during MS and ON.

In silico binding free energy predictability by using the linear interaction energy (LIE) method: bromobenzimidazole CK2 inhibitors as a case study.

Protein kinase CK2 is essential for cell viability, and its control regards a broad series of cellular events such as gene expression, RNA, and protein synthesis. Evidence of its involvement in tumor development and viral replication indicates CK2 as a potential target of antineoplastic and antiviral drugs. In this study the Linear Interaction Energy (LIE) Method with the Surface Generalized Born (SGB) continuum solvation model was used to study several bromobenzimidazole CK2 inhibitors. This methodology, developed by Aqvist, finds a plausible compromise between accuracy and computational speed in evaluating binding free energy (DeltaGbind) values. In this study, two different free binding energy models, named "CK2scoreA" and "CK2scoreB", were developed using 22 inhibitors as the training set in a stepwise approach useful to appropriately select both the tautomeric form and the starting binding position of each inhibitor. Both models are statistically acceptable. Indeed, the better one is characterized by a correlation coefficient (r2) of 0.81, and the predictive accuracy was 0.65 kcal/mol. The corresponding validation, using an external test set of 16 analogs, showed a correlation coefficient (q2) of 0.68 and a prediction root-mean-square error of 0.78 kcal/mol. In this case, the LIE approach has been proved to be an efficient methodology to rationalize the difference of activity, the key interactions, and the different possible binding modes of this specific class of potent CK2 inhibitors.

Cervical-vaginal disease in HIV immunosuppressed patients: management and present screening programme.

The aim of this study was to evaluate the rate of the cervical intraepithelial neoplasia (L-SIL and H-SIL) in HIV-positive patients using cytological, colposcopic and histological examinations. The correlations between these cervical lesions, the role of HPV and the clinical and immunological aspects of HIV infection and inflammatory cervical-vaginal disease were studied. We believe that HPV infection and preneoplastic and/or neoplastic lesions occur more often in immunodepressed HIV-positive patients, and that on the grounds of the high risk of precancerous lesions in this population and the low sensibility of the Pap test, it is advisable to perform a colposcopic examination to discover early lesions that must undergo a specific biopsy.

Novel strategies for the design of new potent and selective human A3 receptor antagonists: an update.

A computer-aided approach has been developed in order to understand the molecular pharmacology of human A3R, and specifically, to lead to the discovery and structural refinement of new, potent and selective human A3R antagonists. This review focuses on our combined target-based and ligand-based drug design strategy, recently applied to provide more accurate information about the recognition mode on human A3R of some pyrazolotriazolopyrimidine and triazoloquinoxalinone analogs. The 3D rhodopsin-based homology model of human A3R has represented the starting point of our approach. A high throughput molecular docking method on the considered antagonists has allowed us to generate a receptor-based pharmacophore model. A novel "Y-shaped" pharmacophore binding motif has been proposed for both pyrazolotriazolopyrimidine and triazoloquinoxalinone derivatives. Moreover, related receptor-based 3D-QSAR analysis has been carried out to provide a suitable tool for prediction of the antagonists binding affinity on human A3R.

Synthesis, biological and modeling studies of 1,3-di-n-propyl-2,4-dioxo-6-methyl-8-(substituted) 1,2,3,4-tetrahydro [1,2,4]-triazolo [3,4-f]-purines as adenosine receptor antagonists.

A new series of potential adenosine receptor antagonists with a [1,2,4]-triazolo-[3,4-f]-purine structure bearing at the 1 and 3 position n-propyl groups have been synthesized, and their affinities at the four human adenosine receptor subtypes (A(1), A(2A), A(2B) and A(3)) have been evaluated. In this case the presence of n-propyl groups seems to induce potency at the A(2A) and A(3) adenosine receptor subtypes as opposed to our previously reported series bearing methyl substituents at the 1 and 3 positions. In particular the non-acylated derivative 17 showed affinity at these two receptor subtypes in the micromolar range. Indeed, preliminary molecular modeling investigations according to the experimental binding data indicate a modest steric and electrostatic antagonist-receptor complementarity.

Discovery of HIV-1 integrase inhibitors through a novel combination of ligand and structure-based drug design.

Over the past 10 years, classical computer-aided molecular design methods have not been frequently applied for the discovery of novel HIV-1 integrase (IN) inhibitors, due to the intrinsic challenges that this enzyme presents. Therefore, a novel approach that combines the chemical information of known integrase inhibitors with the enzyme's detailed 3D structure in a stepwise fashion is proposed: (I) use of a pharmacophore model (PM), which takes into account in a weighted fashion the chemical features of known ligands, in analogous manner to the to search the Maybridge and the NCI 3D databases; (II) drug-likeness optimization; (III) virtual high-throughput screening of the hits matching the PM query against 1QS4 wild-type IN structure using different Docking/Scoring combinations; (IV) visual inspection and selection of the hits in function of: binding free energies; binding mode type within the active site; retrieval among the best 20% hits in more than 6 Docking/Scoring protocols at the same time. This approach aims at a rational selection of new potential HIV-1 integrase inhibitors.

Comparison of pupil responses to luminance and colour in severe optic neuritis.

OBJECTIVE: The pupil response to light flux increments is abnormal in severe optic neuritis, but little is known about the effects of this condition on the pupil colour response. The aim of this study was to examine how optic neuritis affects pupil responses to light flux and colour modulation and the extent to which such pupil responses mirror the loss and recovery of vision. METHODS: A new pupil examination technique that makes use of sinusoidal modulation of either luminance contrast or chromatic saturation was employed. This technique enables the automatic extraction of both pupil response amplitude and latency and achieves a high signal to noise ratio with fewer averages. RESULTS: The study reveals a greater loss of pupil response amplitude and significantly longer latencies to chromatic modulation (i.e. approximately 80 ms). Stimulation of the unaffected eye in the optic neuritis group results in smaller response amplitudes when compared to the normal group for both light flux and colour modulation. CONCLUSIONS: Pupil response components can be affected differently in optic neuritis. These findings suggest that the pupil colour response, in particular, may provide a useful, objective estimator to judge the extent of damage and recovery in diseases of the optic nerve.

PEG-Ara-C conjugates for controlled release.

The antitumour agent 1-beta-D arabinofuranosilcytosyne (Ara-C) was covalently linked to poly(ethylene glycol) (PEG) in order to improve the in vivo stability and blood residence time. Eight PEG conjugates were synthesised, with linear or branched PEG of 5000, 10000 and 20000 Da molecular weight through an amino acid spacer. Starting from mPEG-OH or HO-PEG-OH, conjugation was carried out to the one or two available hydroxyl groups at the polymer's extreme. Furthermore, to increase the drug loading of the polymer, the hydroxyl functions of PEG were functionalised with a bicarboxylic amino acid yielding a tetrafunctional derivative and, by recursive conjugation with the same bicarboxylic amino acid, products with four or eight Ara-C molecules for each PEG chain were prepared. A computer graphic investigation demonstrated that aminoadipic acid was a suitable bicarboxylic amino acid to overcome the steric hindrance between the vicinal Ara-C molecules in the dendrimeric structure. In this paper we report the optimised conditions for synthesis and purification of PEG-Ara-C products with a low amount of remaining free drug, studies toward the hydrolysis of PEG-Ara-C and the Ara-C deamination by cytidine deaminase, pharmacokinetics in mice and cytotoxicity towards HeLa human cells were also investigated. Increased stability towards degradation of the conjugated Ara-C products, in particular for the highly loaded ones, improved blood residence time in mice and a reduced cytotoxicity with respect to the free Ara-C form was demonstrated.

[Bilateral synovial chondromatosis of the first metatarsophalangeal joint: a case report. ]. / Su di un caso di condromatosi sinoviale bilaterale della prima metatarso-falangea.

Synovial chondromatosis is a rare pathology of unknown aetiology. It originates from the chondroid metaplasia of the connective tissue of the synovial membrane. Consequently, cartilaginous nodules develop in the affected joints, first calcifying and then ossifying. The bursae mucosae, the vaginae tendinis and the para-articular connective tissue are less frequently affected. The most common locations of this pathology are the knee, the hip, the shoulder, the elbow and the ankle. The small joints are rarely affected, even less the bilateral involving of joints, above all of hand or foot, is exceptional. In a clinical and radiological valuation, it is difficult to distinguish synovial chondromatosis from osteoarthritis and from degenerative arthopathies in general. A sure diagnosis can be obtained only by means of a histological examination. We here report a case of synovial chondromatosis bilaterally located on the first metatarsophalangeal joint. Clinical and radiological features were similar to those of hallux rigidus, a typical and peculiar metatarsophalangeal joint pathology. The diagnostic suspicion arose during surgery, and was subsequently confirmed by histological examination. During the following visits, the patient did not present any painful symptomatology.

The topoisomerase II poison clerocidin alkylates non-paired guanines of DNA: implications for irreversible stimulation of DNA cleavage.

Clerocidin, a diterpenoid with antibacterial and antitumor activity, stimulates in vitro DNA cleavage mediated by mammalian and bacterial topoisomerase (topo) II. Different from the classical topoisomerase poisons, clerocidin-stimulated breaks at guanines immediately preceding the sites of DNA cleavage are not resealed upon heat or salt treatment. To understand the mechanism of irreversible trapping of the topo II-cleavable complex, we have investigated the reactivity of clerocidin per se towards DNA. We show here that the drug is able to nick negatively supercoiled plasmids. DNA cleavage by clerocidin in enzyme-free medium is due to the ability of the drug to form covalent adducts with guanines. Indeed, clerocidin was able to specifically react with short oligonucleotides when the guanines were unpaired and exposed as in bulges or in the single-strand form. The clerocidin epoxy group attacks the nitrogen at position 7 of guanines, leading to strand scission at the modified site. Our findings also demonstrate that trapping of topoisomerases by clerocidin is specific for type II enzymes. The guanine-alkylating ability of clerocidin suggests an unprecedented mechanism of topo II poisoning, according to which the enzyme renders the drug reactive toward DNA by distorting the double-helical structure of the nucleic acid at the cleavage site.